A viral pan-end RNA element and host complex define a SARS-CoV-2 regulon

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, generates multiple protein-coding, subgenomic RNAs (sgRNAs) from a longer genomic RNA, all bearing identical termini with poorly understood roles in regulating viral gene expression. Insulin and interferon-gamma, two host-derived, stress-related agents, and virus spike protein, induce binding of glutamyl-prolyl-tRNA synthetase (EPRS1), within an unconventional, tetra-aminoacyl-tRNA synthetase complex, to the sgRNA 3′-end thereby enhancing sgRNA expression. We identify an EPRS1-binding sarbecoviral pan-end activating RNA (SPEAR) element in the 3′-end of viral RNAs driving agonist-induction. Translation of another co-terminal 3′-end feature, ORF10, is necessary for SPEAR-mediated induction, independent of Orf10 protein expression. The SPEAR element enhances viral programmed ribosomal frameshifting, thereby expanding its functionality. By co-opting noncanonical activities of a family of essential host proteins, the virus establishes a post-transcriptional regulon stimulating global viral RNA translation. A SPEAR-targeting strategy markedly reduces SARS-CoV-2 titer, suggesting a pan-sarbecoviral therapeutic modality.


March 2021
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Replication Randomization spectrometry-lc-ms/lc-ms-software/multi-omics-data-analysis/proteome-discoverer-software.html) Genscript Rare Codon Analysis Tool (https://www.genscript.com/tools/rare-codon-analysis) Rare Codon Analyzer (https://www.biologicscorp.com/tools/RareCodonAnalyzer/#.Y_7PGXbMK70) Further information and requests for resources and reagents should be directed to, and will be fulfilled by, the Lead Contact, Paul L. Fox (foxp@ccf.org). Requests for SARS-CoV-2 reporter virus should be directed to, and will be fulfilled by, Michaela U. Gack (gackm@ccf.org). Requests for SARS-CoV2N-EGFP replicon can be directed to, and will be fulfilled by, Dr. Jonathan Karn (jxk153@case.edu). All stable reagents generated in this study are available from the Lead Contacts without restriction, or with a Materials Transfer Agreement. The dORF8-EGFP rSARS-CoV-2 will require an MTA. Source data are provided with this paper. All graph data used in this study are available in the accompanying Source Data file. All raw micrographs used in this study are available in the accompanying Source Data file. Data points were not excluded.
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Antibodies validated with knockdown (siRNA/shRNA) or knockout approach by seller, in published studies or during this study, except LARS1, IARS1, SARS1 and EEF1A, that were validated here by molecular weight according to SDS-PAGE. Anti-KDEL antibody detects a group of endoplasmic reticulum-targeted proteins and in absence of any one specific target, is tested by enrichment in biochemically isolated subcellular microsomal fraction. Anti-Nsp3, spike, Orf3a, M, Orf7a and N antibodies were validated by molecular weight in SDS-PAGE as well as presence of signal specifically in SARS-CoV-2 infected cells over uninfected cells. Available application-specific and species-specific testing and validation information from seller websites have been added in Supplementary Table 4 under each antibody catalog number.